Setting the stage for universal pharmacological targeting of the glycocalyx.

All cells in the human body are covered by a complex meshwork of sugars as well as proteins and lipids to which these sugars are attached, collectively termed the glycocalyx. Over the past few decades, the glycocalyx has been implicated in a range of vital cellular processes in health and disease. Therefore, it has attracted considerable interest as a therapeutic target. Considering its omnipresence and its relevance for various areas of cell biology, the glycocalyx should be a versatile platform for therapeutic intervention, however, the full potential of the glycocalyx as therapeutic target is yet to unfold. This might be attributable to the fact that glycocalyx alterations are currently discussed mainly in the context of specific diseases. In this perspective review, we shift the attention away from a disease-centered view of the glycocalyx, focusing on changes in glycocalyx state. Furthermore, we survey important glycocalyx-targeted drugs currently available and finally discuss future steps. We hope that this approach will inspire a unified, holistic view of the glycocalyx in disease, helping to stimulate novel glycocalyx-targeted therapy strategies.

Oscillators in the microvasculature: glycocalyx and beyond.

The growing recognition of abundance of oscillating functions in biological systems has motivated this brief overview, which narrows down on the microvasculature. Specifically, it encompasses self-sustained oscillations of blood flow, hematocrit, and viscosity at bifurcations; blood flow effects on the oscillations of endothelial glycocalyx, mechanotransduction, and its termination to prime endothelial cells for the subsequent mechanical signaling event; oscillating affinity of hyaluronan-CD44 binding domain; spontaneous contractility of actomyosin complexes in the cortical actin web and its effects on the tension of the plasma membrane; reversible effects of sirtuin-1 on endothelial glycocalyx; and effects of plasma membrane tension on endo- and exocytosis. Some potential interactions between those oscillators, and their coupling, are discussed together with their transition into chaotic movements. Future in-depth understanding of the oscillatory activities in the microvasculature could serve as a guide to its chronotherapy under pathological conditions.

Anti-inflammatory effects of a SERP 30 polysaccharide from the residue of Sarcandra glabra against lipopolysaccharide-induced acute respiratory distress syndrome in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Sarcandra glabra (Thunb.) Nakai, a valuable dietetic Chinese herb, is still widely used today. Multiple ingredients of S. glabra with a variety of activities such as anti-inflammatory, antiviral, and antitumor were studied. However, the Sarcandra glabra (Thunb.) Nakai polysaccharide hasn't been reported for its anti-inflammatory effect. AIM OF THE STUDY: In this study, the anti-inflammatory activity of Sarcandra glabra (Thunb.) Nakai polysaccharide was assessed in LPS-induced ARDS mice. MATERIALS AND METHODS: A polysaccharide coded as SERP 30 was obtained by water extraction, alcohol precipitation, and gel filtration. After the physicochemical properties determination and structural characterization, LPS induced-mice ARDS model was used to evaluate the anti-inflammatory and associated antioxidant activities of SERP 30. H&E staining was used to observe the seriousness of lung injury in mice. The ELISA method was used to measure the expression of inflammatory factors (TNF-α and IL-6) in the serum of the mice. The TBA method and the WST-1 method were used to evaluate the oxidative stress injury. Immunohistochemistry was used to distinguish the expression of metalloproteinase-9 (MMP-9), heparinase (HPA), syndecan-1, and decorin in ARDS-mice lung tissue. Western blotting was used to confirm the expression of related proteins in mouse lung tissue. RESULTS: SERP 30 had a potential role in improving lung damage, reducing inflammation, and preventing oxidative stress. Moreover, SERP 30 significantly attenuated the damage to the endothelial glycocalyx and maintained the integrity of the glycocalyx. The western blotting result implied that the main anti-inflammatory mechanism is directed towards NF-κB and MAPK signaling pathways with inhibiting the activation of associated proteins. CONCLUSION: This research provides a theoretical basis for treating ARDS by using a byproduct from food resource.

Roles of glomerular endothelial hyaluronan in the development of proteinuria.

Vascular endothelial cells are covered with glycocalyx comprising heparan sulfate, hyaluronan, chondroitin sulfate, and associated proteins. Glomerular endothelial glycocalyx is involved in protecting against induction of proteinuria and structural damage, but the specific components in glycocalyx that represent therapeutic targets remain unclear. Anti-vascular endothelial growth factor (VEGF) therapy is associated with an increased risk of glomerular endothelial injury. This study investigated whether hyaluronan could provide a therapeutic target to protect against proteinuria. We conducted ex vivo and in vivo experiments to explore the effects of degrading glomerular hyaluronan by administering hyaluronidase and of supplementation with hyaluronan. We investigated hyaluronan expression using biotin-labeled hyaluronan-binding protein (HABP) in human kidney specimens or serum hyaluronan in endothelial injuries under inhibition of VEGF signaling. We directly demonstrated hyaluronan in glomerular endothelial layers using HABP staining. Ex vivo and in vivo experiments showed the development of proteinuria after digestion of hyaluronan in glomerular capillaries. Supplementation with hyaluronan after hyaluronidase treatment suppressed proteinuria. Mice in the in vivo study developed albuminuria after intraperitoneal injection of hyaluronidase with decreased glomerular hyaluronan and increased serum hyaluronan. In human kidneys with endothelial cell dysfunction and proteinuria due to inhibition of VEGF, glomerular expression of hyaluronan was reduced even in normal-appearing glomeruli. Serum hyaluronan levels were elevated in patients with pre-eclampsia with VEGF signaling inhibition. Our data suggest that hyaluronan itself plays crucial roles in preventing proteinuria and preserving the integrity of endothelial cells. Hyaluronan could provide a therapeutic target for preventing glomerular endothelial glycocalyx damage, including VEGF signaling inhibition.

In vitro evaluation of different organic matrices used to modulate silicon bioavailability.

Silicon (Si) has numerous health properties. It is an element of the extracellular matrix; it is involved in collagen synthesis, bone mineralization, and immune system modulation; and it reduces metal accumulation in Alzheimer's disease and the risk of atherosclerosis. Given its poor intestinal absorption, Si is ingested in the form of orthosilicic acid (OSA) to promote its bioavailability. The aim of this work was to compare different commercial dietary supplements containing stabilized OSA to ascertain their bioaccessibility, bioavailability, and safety in a model of human intestinal epithelium. Biocompatibility with the glycocalyx was also investigated. Supplements containing collagen, maltodextrins, and choline as OSA stabilizers were analyzed. Bioaccessibility was explored by means of an in vitro digestive process. Bioavailability was investigated using a Caco2 cell line alone, or co-culturing with a HT29-MTX cell line. The safety of the compounds tested (in terms of intestinal epithelium integrity) was judged on the grounds of MTS assay, transepithelial electrical resistance, and apparent permeability. The three formulations were also tested in a Caco2 cell model of intestinal glycocalyx Si retention. The choline-formulated OSA formulation outperformed the maltodextrin-stabilized supplement, with a Si bioavailability about 14 times higher (P < .05). The choline-formulated OSA formulation increased cell permeability, with consequent intestinal epithelium disruption. The supplements' absorption and bioavailability (and harmfulness) differed considerably, depending on the OSA stabilizer involved. Of the three formulations tested, the collagen-formulated OSA represents the best Si dietary supplement.

Modulation of Endothelial Glycocalyx and Microcirculation in Healthy Young Men during High-Intensity Sprint Interval Cycling-Exercise by Supplementation with Pomegranate Extract. A Randomized Controlled Trial.

The natural components of the pomegranate fruit may provide additional benefits for endothelial function and microcirculation. It was hypothesized that supplementation with pomegranate extract might improve glycocalyx properties and microcirculation during acute high-intensity sprint interval cycling exercise. Eighteen healthy and recreationally active male volunteers 22-28 years of age were recruited randomly to the experimental and control groups. The experimental group was supplemented with pomegranate extract 20 mL (720 mg phenolic compounds) for two weeks. At the beginning and end of the study, the participants completed a high-intensity sprint interval cycling-exercise protocol. The microcirculation flow and density parameters, glycocalyx markers, systemic hemodynamics, lactate, and glucose concentration were evaluated before and after the initial and repeated (after 2 weeks supplementation) exercise bouts. There were no significant differences in the microcirculation or glycocalyx over the course of the study (p < 0.05). The lactate concentration was significantly higher in both groups after the initial and repeated exercise bouts, and were significantly higher in the experimental group compared to the control group after the repeated bout: 13.2 (11.9-14.8) vs. 10.3 (9.3-12.7) mmol/L, p = 0.017. Two weeks of supplementation with pomegranate extract does not influence changes in the microcirculation and glycocalyx during acute high-intensity sprint interval cycling-exercise. Although an unexplained rise in blood lactate concentration was observed.

Goal-Directed Fluid Therapy Does Not Improve Early Glomerular Filtration Rate in a Porcine Renal Transplantation Model.

BACKGROUND: Insufficient fluid administration intra- and postoperatively may lead to delayed renal graft function (DGF), while fluid overload increases the risk of heart failure, infection, and obstipation. Several different fluid protocols have been suggested to ensure optimal fluid state. However, there is a lack of evidence of the clinical impact of these regimens. This study aimed to determine whether individualized goal-directed fluid therapy (IGDT) positively affects the initial renal function compared to a high-volume fluid therapy (HVFT) and to examine the effects on renal endothelial glycocalyx, inflammatory and oxidative stress markers, and medullary tissue oxygenation. The hypothesis was that IGDT improves early glomerular filtration rate (GFR) in pigs subjected to renal transplantation. METHODS: This was an experimental randomized study. Using a porcine renal transplantation model, animals were randomly assigned to receive IGDT or HVFT during and until 1 hour after transplantation from brain-dead donors. The kidneys were exposed to 18 hours of cold ischemia. The recipients were observed until 10 hours after reperfusion, which included GFR measured as clearance of chrom-51-ethylendiamintetraacetat (Cr-EDTA), animal weight, and renal tissue oxygenation by fiber optic probes. The renal expression of inflammatory and oxidative stress markers as well as glomerular endothelial glycocalyx were analyzed in the graft using polymerase chain reaction (PCR) technique and immunofluorescence. RESULTS: Twenty-eight recipient pigs were included for analysis. We found no evidence that IGDT improved early GFR compared to HVFT (P = .45), while animal weight increased more in the HVFT group (a mean difference of 3.4 kg [1.96-4.90]; P < .0001). A better, however nonsignificant, preservation of glomerular glycocalyx (P = .098) and significantly lower levels of the inflammatory marker cyclooxygenase 2 (COX-2) was observed in the IGDT group when compared to HVFT. COX-2 was 1.94 (1.50-2.39; P = .012) times greater in the HVFT group when compared to the IGDT group. No differences were observed in outer medullary tissue oxygenation or oxidative stress markers. CONCLUSIONS: IGDT did not improve early GFR; however, it may reduce tissue inflammation and could possibly lead to preservation of the glycocalyx compared to HVFT.

Fraxin Alleviates LPS-Induced ARDS by Downregulating Inflammatory Responses and Oxidative Damages and Reducing Pulmonary Vascular Permeability.

Acute respiratory distress syndrome (ARDS) is a severe acute disease that threatens human health, and few drugs that can effectively treat this disease are available. Fraxin, one of the main active ingredients of Cortex Fraxini, a Chinese herbal medicine, has presented various pharmacological and biological activities. However, the effects of fraxin on ARDS have yet to be reported. In the present study, the protective effect of fraxin in lipopolysaccharide (LPS)-induced ARDS in a mouse model was analyzed. Results from the hematoxylin and eosin staining showed that fraxin might alleviate pathological changes in the lung tissues of mice with ARDS. ELISA and Western blot results revealed that fraxin might inhibit the production of inflammatory factors, namely, IL-6, TNF-α, and IL-1ß, and the activation of NF-κB and MAPK signaling pathways in the lungs. Thus, the inflammatory responses were reduced. Fraxin might inhibit the increase in reactive oxygen species (ROS) and malondialdehyde (MDA), a product of lipid peroxidation in lung tissues. Fraxin might increase the superoxide dismutase (SOD) activity to avoid oxidative damage. Vascular permeability was also assessed through Evans blue dye tissue extravasation and fluorescein isothiocyanate-labeled albumin (FITC-albumin) leakage. Fraxin might inhibit the increase in pulmonary vascular permeability and relieve pulmonary edema. Fraxin was also related to the inhibition of the increase in matrix metalloproteinase-9, which is a glycocalyx-degrading enzyme, and the relief of damages to the endothelial glycocalyx. Thus, fraxin elicited protective effects on mice with LPS-induced ARDS and might be used as a drug to cure ARDS induced by Gram-negative bacterial infection.

Structural heteropolysaccharides as air-tight sealants of the human pleura.

Pulmonary "air leaks," typically the result of pleural injury caused by lung surgery or chest trauma, result in the accumulation of air in the pleural space (pneumothorax). Air leaks are a major source of morbidity and prolonged hospitalization after pulmonary surgery. Previous work has demonstrated structural heteropolysaccharide (pectin) binding to the mouse pleural glycocalyx. The similar lectin-binding characteristics and ultrastructural features of the human and mouse pleural glycocalyx suggested the potential application of these polymers in humans. To investigate the utility of pectin-based polymers, we developed a simulacrum using freshly obtained human pleura. Pressure-decay leak testing was performed with an inflation maneuver that involved a 3 s ramp to a 3 s plateau pressure; the inflation was completely abrogated after needle perforation of the pleura. Using nonbiologic materials, pressure-decay leak testing demonstrated an exponential decay with a plateau phase in materials with a Young's modulus less than 5. In human pleural testing, the simulacrum was used to test the sealant function of four mixtures of pectin-based polymers. A 50% high-methoxyl pectin and 50% carboxymethylcellulose mixture demonstrated no sealant failures at transpleural pressures of 60 cmH2 O. In contrast, pectin mixtures containing 50% low-methoxyl pectin, 50% amidated low-methoxyl pectins, or 100% carboxymethylcellulose demonstrated frequent sealant failures at transpleural pressures of 40-50 cmH2 O (p < 0.001). Inhibition of sealant adhesion with enzyme treatment, dessication and 4°C cooling suggested an adhesion mechanism dependent upon polysaccharide interpenetration. We conclude that pectin-based heteropolysaccharides are a promising air-tight sealant of human pleural injuries. © 2018 Wiley Periodicals, Inc. J. Biomed. Mater. Res. Part B, 2018. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 799-806, 2019.

Glycocalyx bending by an electric field increases cell motility.

The application of physiological strength electric fields may produce a wide range of effects on cells. The mechanisms by which cells detect the presence of these fields, however, are not fully understood. Previous experiments have shown that directionality of cells in the field is governed by an electromechanical mechanism in which the field exerts a torque on the negatively charged, inner glycocalyx that is then transmitted as a force on the cytoskeleton. This mechanism is similar to that by which cells detect fluid shear forces. Several authors, however, have reported that cell directionality and motility behave differently in an electric field. We propose here a second electromechanical mechanism in which the field bends the negatively charged, outer glycocalyx in proximity to the substrate, increasing cell adhesion and, thus, cell motility. The increase in motility depends not only on the field strength, but also on the adhesion of the cell to the substrate prior to application of the field. We show that these mechanisms are common to both human cells and amoebae and, hence, are evolutionarily conserved. Furthermore, the mechanism for detection of electric fields is simply an extension of the mechanism for detecting fluid shears. Bioelectromagnetics. 38:482-493, 2017. © 2017 Wiley Periodicals, Inc.

Anesthetic propofol overdose causes vascular hyperpermeability by reducing endothelial glycocalyx and ATP production.

Prolonged treatment with a large dose of propofol may cause diffuse cellular cytotoxicity; however, the detailed underlying mechanism remains unclear, particularly in vascular endothelial cells. Previous studies showed that a propofol overdose induces endothelial injury and vascular barrier dysfunction. Regarding the important role of endothelial glycocalyx on the maintenance of vascular barrier integrity, we therefore hypothesized that a propofol overdose-induced endothelial barrier dysfunction is caused by impaired endothelial glycocalyx. In vivo, we intraperitoneally injected ICR mice with overdosed propofol, and the results showed that a propofol overdose significantly induced systemic vascular hyperpermeability and reduced the expression of endothelial glycocalyx, syndecan-1, syndecan-4, perlecan mRNA and heparan sulfate (HS) in the vessels of multiple organs. In vitro, a propofol overdose reduced the expression of syndecan-1, syndecan-4, perlecan, glypican-1 mRNA and HS and induced significant decreases in the nicotinamide adenine dinucleotide (NAD+)/NADH ratio and ATP concentrations in human microvascular endothelial cells (HMEC-1). Oligomycin treatment also induced significant decreases in the NAD+/NADH ratio, in ATP concentrations and in syndecan-4, perlecan and glypican-1 mRNA expression in HMEC-1 cells. These results demonstrate that a propofol overdose induces a partially ATP-dependent reduction of endothelial glycocalyx expression and consequently leads to vascular hyperpermeability due to the loss of endothelial barrier functions.

Sodium selective erythrocyte glycocalyx and salt sensitivity in man.

Negatively charged surfaces of erythrocytes (RBC) reflect properties of the endothelial glycocalyx. Plasma electrolytes counteract these charges and thus control the repulsive forces between RBC and endothelium. Although Na(+) is supposed to exert a rather high affinity to the RBC surface, a direct comparison between Na(+) and K(+) in counteracting the RBC surface has been never made. Therefore, we measured Na(+)/K(+) selectivity of the RBC surface in 20 healthy volunteers applying the previously published salt blood test (SBT). It turned out that the Na(+)/K(+) selectivity ratio of the RBC glycocalyx is on average 6.1 ± 0.39 (ranging from 3 to 9 in different individuals). Considering standard plasma Na(+) and K(+) concentrations, binding probability of Na(+)/K(+) at the RBC surface is about 180:1. The SBT reveals that plasma K(+) counteracts only about 7% of the negative charges in the RBC glycocalyx. As an in vivo proof of principle, a volunteer's blood was continuously tested over 6 months while applying a glycocalyx protective polyphenol-rich natural compound (hawthorn extract). It turned out that RBC Na(+) sensitivity (the inverse of Na(+) buffer capacity) decreased significantly by about 25% while Na(+)/K(+) selectivity of the RBC glycocalyx declined only slightly by about 8 %. Taken together, (i) plasma Na(+) selectively buffers the negative charges of the RBC glycocalyx, (ii) the contribution of K(+) in counteracting these negative surface charges is small, and (iii) natural polyphenols applied in vivo increase RBC surface negativity. In conclusion, low plasma Na(+) is supposed to favor frictionless RBC-slipping through blood vessels.

Sevoflurane preserves the endothelial glycocalyx against ischaemia-reperfusion injury.

BACKGROUND: Healthy vascular endothelium is coated by the glycocalyx, important in multiple endothelial functions, but destroyed by ischaemia-reperfusion. The impact of volatile anaesthetics on this fragile structure has not been investigated. We evaluated the effect of cardiac pre- and post-conditioning with sevoflurane on integrity of the endothelial glycocalyx in conjunction with coronary vascular function. METHODS: Isolated guinea pig hearts perfused with Krebs-Henseleit buffer underwent 20 min stopped-flow ischaemia (37 degrees C), either without or with 1 MAC sevoflurane. This was applied for 15 min before, for 20 min after, or both before and after ischaemia. Transudate was collected for assessing coronary net fluid extravasation and histamine release by mast cells. Coronary release of syndecan-1 and heparan sulphate was measured. In additional experiments with and without continuous sevoflurane, cathepsin B and tryptase beta-like protease activity were measured in effluent. Hearts were perfusion-fixed to visualize the endothelial glycocalyx. RESULTS: Ischaemia led to a significant (P<0.05) increase by 70% in transudate formation during reperfusion only in hearts without sevoflurane. This was accompanied by significant (P<0.05) increases in heparan sulphate (four-fold) and syndecan release (6.5-fold), with electron microscopy revealing massive degradation of glycocalyx. After ischaemia, histamine was released into transudate, and cathepsin B activity increased in effluent (P<0.05). Sevoflurane application attenuated all these changes, except for histamine release. CONCLUSIONS: Sevoflurane protects the endothelial glycocalyx from ischaemia-reperfusion-induced degradation, with both preconditioning and rapid post-conditioning being successful. The mechanism seems to involve attenuation of lysosomal cathepsin B release and to be independent from tissue mast cell degranulation.